Project Summary/Abstract Morphogenesis is the fundamental developmental process that drives tissue assembly and elaborates the diverse anatomical structures that together comprise the body plans of all metazoa. Most human birth defects arise from disruptions in normal morphogenetic processes. How morphogenesis works at multiple levels of organization and complexity is one of the key remaining questions in biology and progress in this area will be needed to help inform efforts to engineer replacement tissues and organs. For nearly three decades our laboratory has approached this problem by focusing on the cell movements and tissue rearrangements responsible for gastrulation in the amphibian Xenopus. We explore how cell adhesion to other cells and to the extracellular matrix (ECM) is regulated to promote or stabilize the cell and tissue movements of gastrulation. Cadherin and integrin adhesion complexes are central players in these processes and aside from their general roles in holding cells and tissues together, they also sense, resist and distribute mechanical forces that arise as a consequence of morphogenesis. We have discovered a novel function for cadherins and keratin intermediate filaments (KIF) in the force-dependent regulation of collective cell migration in the mesendoderm at gastrulation. Although the importance of adherens junctions, focal adhesions and the actin cytoskeleton to mechanosensation and mechanotransduction is now well established, the role of desmosomes and other intermediate filament (IF) associated junctions in these processes has been largely overlooked. A major goal of this grant proposal is to bridge this significant gap in understanding by focusing on KIF functions and the association of the KIF cytoskeleton with cadherin-based adhesions in the mesendoderm. We hypothesize that the magnitude of forces applied to cadherins can direct the differential recruitment and assembly of adhesions linked to the actin or KIF cytoskeletons. We will test this by developing new assays designed to apply defined forces to cadherins on single cells and follow the recruitment of adherens- and desmosomal-associated junctional proteins to stressed adhesion sites. In related experiments we will also ask whether patterning of mesoderm genes varies with the magnitude of forces applied to cell-cell and/or cell-matrix adhesions, perhaps acting analogously to (and/or in synergy with) cells sensing positional information within a concentration gradient of morphogen. Future progress in these areas will also benefit from the development of in silico simulations of morphogenetic movements. A team of collaborators expert in agent-based and finite element methods for modelling cell and tissue-level behaviors will work with us to simulate initially the force dependent polarization and collective migration of mesendoderm cells. Our longer-term goal is to explore ways to integrate simulations of multiple regional morphogenetic machines to gain a better picture of the global contributions of individual tissue movements to gastrulation.